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| Status |
Public on Oct 23, 2014 |
| Title |
Genome-wide RNA tomography in the zebrafish embryo |
| Organisms |
Danio rerio; Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
Advancing our understanding of embryonic development is heavily dependent on identification of novel pathways or regulators. While genome-wide techniques such as RNA sequencing are ideally suited for discovering novel candidate genes, they are unable to yield spatially resolved information in embryos or tissues. Microscopy-based approaches, using for example in situ hybridization, can provide spatial information about gene expression, but are limited to analyzing one or a few genes at a time. Here, we present a method where we combine traditional histological techniques with low-input RNA sequencing and mathematical image reconstruction to generate a high-resolution genome-wide 3D atlas of gene expression in the zebrafish embryo at three developmental stages. We also demonstrate that our technique is suitable for spatially-resolved differential expression analysis in wildtype and Gli3 mutant mouse forelimbs. Importantly, our method enables searching for genes that are expressed in specific spatial patterns without manual image annotation. We envision broad applicability of RNA tomography as an accurate and sensitive approach for spatially resolved transcriptomics in whole embryos and dissected organs.
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| Overall design |
To generate spatially-resolved RNA-seq data for zebrafish embryos (shield stage, 10 somites, 15 somites, 18 somites) and mouse forelimbs (E10.5), we cryosectioned samples, extracted RNA from the individual sections, and amplified and barcoded mRNA using the CEL-seq protocol (Hashimshony et al., Cell Reports, 2012) with a few modifications. Libraries were sequenced on Illumina HiSeq 2500 using 50bp paired end sequencing. Selected zebrafish libraries were sequenced on MiSeq 250bp paired-end to improve 3' annotations.
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| Web link |
http://zebrafish.genomes.nl/tomoseq/
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| Contributor(s) |
Junker JP, Noël ES, Guryev V, Peterson KA, Shah G, Huisken J, McMahon AP, Berezikov E, Bakkers J, van Oudenaarden A |
| Citation(s) |
25417113 |
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| Submission date |
Jul 29, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Jan Philipp Junker |
| Organization name |
Hubrecht Institute
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| Street address |
Uppsalalaan 8
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| City |
Utrecht |
| ZIP/Postal code |
3584CT |
| Country |
Netherlands |
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| Platforms (2) |
| GPL17021 |
Illumina HiSeq 2500 (Mus musculus) |
| GPL18413 |
Illumina HiSeq 2500 (Danio rerio) |
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| Samples (34)
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| GSM1448795 |
shield stage, sectioned from animal to vegetal pole |
| GSM1448796 |
shield stage, sectioned from vegetal to animal pole |
| GSM1448797 |
shield stage, sectioned from vegetal to animal pole, replicate |
| GSM1448798 |
shield stage, sectioned from left to right |
| GSM1448799 |
shield stage, sectioned from right to left |
| GSM1448800 |
shield stage, sectioned from ventral to dorsal end |
| GSM1448801 |
shield stage, sectioned from dorsal to ventral end |
| GSM1448802 |
shield stage, sectioned from dorsal to ventral end, replicate |
| GSM1448803 |
10 somite stage, sectioned from anterior to posterior end |
| GSM1448804 |
10 somite stage, sectioned from anterior to posterior end, replicate |
| GSM1448805 |
10 somite stage, sectioned from posterior to anterior end |
| GSM1448806 |
10 somite stage, sectioned from ventral to dorsal end |
| GSM1448807 |
10 somite stage, sectioned from dorsal to ventral end |
| GSM1448808 |
10 somite stage, sectioned from left to right |
| GSM1448809 |
10 somite stage, sectioned from right to left |
| GSM1448810 |
15 somite stage, sectioned from anterior to posterior end |
| GSM1448811 |
15 somite stage, sectioned from posterior to anterior end |
| GSM1448812 |
15 somite stage, sectioned from ventral to dorsal end |
| GSM1448813 |
15 somite stage, sectioned from dorsal to ventral end |
| GSM1448814 |
15 somite stage, sectioned from left to right |
| GSM1448815 |
15 somite stage, sectioned from right to left |
| GSM1448816 |
15 somites stage, straightened, sectioned head to tail |
| GSM1448817 |
15 somites stage, straightened, sectioned tail to head |
| GSM1448818 |
sections 97-108 for samples dr_15ss_TH and dr_15ss_HT |
| GSM1448819 |
15 somites stage, straightened, sectioned tail to head, replicate |
| GSM1448820 |
18 somites stage, straightened, sectioned tail to head |
| GSM1448821 |
forelimb, wildtype, high resolution |
| GSM1448822 |
forelimb, wildtype, low resolution, sample 1 |
| GSM1448823 |
forelimb, wildtype, low resolution, sample 2 |
| GSM1448824 |
forelimb, wildtype, low resolution, sample 3 |
| GSM1448825 |
forelimb, Gli3-/- high resolution |
| GSM1448826 |
forelimb, Gli3-/-, low resolution, sample 1 |
| GSM1448827 |
forelimb, Gli3-/-, low resolution, sample 2 |
| GSM1448828 |
forelimb, Gli3-/-, low resolution, sample 3 |
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| Relations |
| BioProject |
PRJNA256975 |
| SRA |
SRP045064 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE59873_RAW.tar |
19.6 Mb |
(http)(custom) |
TAR (of TXT) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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